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  • Ma, Hairan (VTT Technical Research Centre of Finland Ltd, 2015)
    Milk proteins, sodium caseinate (CN) and whey protein isolate (WPI) are used in food industries as emulsifiers. The stability of an O/W emulsion is dependent on the electrostatic and steric repulsion provided by the interfacial proteins against droplet aggregation or lipid oxidation. Therefore, modifications of the surface charge or the interfacial conformation of protein emulsifiers are expected to enhance their emulsifying properties and emulsion stability. In this present work, sodium caseinate and whey protein isolate were modified by different chemical and enzymatic approaches. The modified proteins were characterized using multiple techniques, and the effect of these modifications on emulsifying properties of proteins and emulsion stability was investigated. Succinylation converts the positively charged amino groups into negatively charged carboxyl groups, lowering the isoelectric point (pI) of protein. The ethylene diamine (EDA) modification worked in the opposite way, leading to an increased pI. The extent of these two modifications was studied using SDS-PAGE and MALDI-TOF mass spectrometry. The pI of succinylated and EDA modified milk proteins was studied using zeta-potential measurement. As a result, the succinylation to full extent altered the pI of CN from 4.2 to 2.7, and the EDA modification shifted the pI of CN and WPI from 4.2 to 9.4 and from 4.9 to 9.5 respectively. The pH stability of emulsion made with the modified milk proteins was monitored by following the increase of particle size during storage. The results suggested that succinylation and EDA modification could enhance the emulsion stability at pH 4 7 by increasing the electrostatic repulsion between droplets. Regarding the enzymatic modification of milk proteins, the laccase and transglutaminase (Tgase) catalyzed cross-linking were applied on WPI and CN respectively. In order to improve the reactivity of WPI towards the laccase, a vanillic acid modification was carried out to incorporate additional methoxyphenol groups into the protein surface. The cross-linking of vanillic acid modified WPI (Van-WPI) by laccase was studied using SDS-PAGE. The extent of cross-linking of Van-WPI was found to be significantly higher compared to the unmodified WPI and the combination of WPI and free phenolic compound as a mediator. The effect of laccase catalyzed cross-linking on storage stability of emulsions was investigated by visual observation and confocal microscopy. The post-emulsification cross-linking was proven to enhance the stability of the emulsions prepared with Van-WPI during the storage. The reduced droplet coalescence could be most likely attributed to an extended interfacial protein layer formed via the interaction between the adsorbed proteins and non-adsorbed proteins in the water phase. In contrast with the limited extent of cross-linking of WPI by laccase, CN was extensively cross-linked by Tgase. The physical stability of emulsions was studied by measuring the increase of particle size during storage, and the oxidative stability was evaluated by following the formation of fatty acid hydroperoxides and volatile compounds in different stages of the lipid oxidation. The pre-emulsification cross-linking showed no obvious influence on the physical stability of CN emulsion but significantly improved its stability against lipid oxidation. The improvement of oxidative stability of emulsions could be contributed to a thicker and denser interfacial protein layer, which thus increases the amount of anti-oxidative groups located at the interface and provides a stronger barrier against competitive adsorption by oil oxidation products.
  • Knaster, Peter (Helsingin yliopisto, 2015)
    Chronic pain is a long-lasting burdensome condition. Comorbid psychological symptoms are common in chronic pain patients and they tend to worsen the treatment outcome. The aim of this cross-sectional study was to assess the prevalence of psychiatric comorbidity in chronic pain patients and to assess the associations between chronic pain, anxiety, anger, and depression. The study participants in the study were one hundred consecutive chronic pain patients referred to the Meilahti Pain Clinic of Helsinki University Hospital. The subjects were interviewed with the Structured Clinical Interview for DSM-IV for Axis I disorders (SCID-I). In addition self-report questionnaires were used in the assessment. Most (75%) patients had at least one lifetime mental disorder. The prevalence of major depressive disorder (MDD) was 37% and of a specific anxiety disorder 25% over the past 12 months. The psychiatric comorbidity was associated with increased pain intensity, measured by Visual Analogue Scale (VAS). The majority (77%) of the anxiety disorders had their onset before the onset of pain, whereas only 37% of the mood disorders preceded pain onset. The Harm Avoidance (HA) scale of the Temperament and Character Inventory (TCI) of Cloninger was associated with pain-related anxiety measured with Pain Anxiety Symptom Scale-20. The pain intensity influenced the strength of the association between the Harm Avoidance HA4 subscale and pain-related anxiety. Higher pain intensity was associated with stronger association between the variables. Likewise, pain intensity influenced the association between anger management and depression. A tendency to inhibit angry feelings was associated the somatic and physical symptoms of depression, measured by a two-factor model of the Beck Depression Inventory (BDI). The association was stronger in patients with higher pain intensity. Chronic pain patients with current MDD scored higher in both the somatic and cognitive-emotional subscales of the BDI compared with those without MDD. However, the somatic-physical-related items were more strongly associated with the diagnosis of MDD. Psychiatric disorders are common in chronic pain patients. Because of their influence on the chronic pain treatment and outcome, their thorough assessment is important. The linking mechanisms between chronic pain and the psychiatric disorders and symptoms are complicated and there is an overlap between the constructs. Clear boundaries between pain, anxiety and depression can be difficult to draw, which can be a reflection of the common background mechanisms such as common neural circuits and neurotransmitters.
  • Morandin, Claire (Helsingin yliopisto, 2015)
    Understanding how the interaction of genotypes and environment may result in distinct phenotypes from similar sets of genes is a central theme in evolutionary biology. Social insects have been important study organisms for this question, with sociality evolving in parallel in unrelated taxa. The defining feature of advanced sociality, the separate reproductive queen and non-reproductive worker castes of social insects, is central to social evolution. Queens and workers share, most of the time, a similar genome, suggesting that the basis of this dimorphism must result from differences in expression of the same genes. Furthermore, workers forgo their own reproduction to help raise the offspring of the queen, thus queen and worker genes are expected to experience natural selection in unique ways. Yet, the regulatory architecture that governs queen and worker phenotypes remains largely unknown in social insects. This thesis demonstrates the plasticity of caste-biased expression patterns in ants at several levels. It shows that few genes retain their caste-biased expression patterns across closely related species, lineages, or development stages. For the first time, this thesis shows that biological functions, through conserved sets of genes, are strongly associated with caste phenotypic differences across the ant phylogeny. These sets of genes also appear to be co-opted for other types of key social phenotypes and likely serve as building blocks of phenotypic innovation. My thesis furthermore focuses on the interactions between caste-biased expression patterns and rates of molecular evolution to comprehend the origins and results of caste-biases. The work presented in this thesis shows that evolutionary constraints strongly affect evolutionary rates of protein-coding genes, gene expression evolutionary stability, and the ability of a gene to become caste-biased. These constraints are significant features that have been greatly under-appreciated in previous studies. The work in this thesis takes advantage of the power of genomic methodologies and technologies to provide new insights into mechanisms of social evolution, and the evolution of plastic gene expression in a more general framework. It builds on existing knowledge to provide the field of social insect research with novel concepts (e.g. co-expressed network) to understand the molecular mechanisms behind the origin and the maintenance of the two female castes.
  • Tumiati, Manuela (Helsingin yliopisto, 2015)
    Breast cancer is the second most common cancer in the world and the most common cancer among women. Germ-line mutations in the DNA repair gene RAD51C (RAD51 paralog C) predispose women to breast and ovarian cancers, yet the mechanisms by which a lack of RAD51C causes tumorigenesis are poorly understood. RAD51C deficiency is thought to promote cancer by preventing correct repair of DNA double-strand breaks, leading to accumulation of somatic mutations and genomic instability, a cancer hallmark. Similarly, defects in other genes involved in repair of DNA double-strand breaks, such as BRCA1 (breast cancer 1, early onset), BRCA2 (breast cancer 2, early onset), or PALB2 (partner and localizer of BRCA2), are linked to breast cancer, suggesting that the mammary gland is particularly susceptible to genomic instability. We know that RAD51C-null cells from several organisms present a number of chromosomal aberrations, and Rad51c knockout mice die during early embryogenesis from massive Trp53-mediated apoptosis. A previously generated mouse model demonstrated that when Rad51c is lost together with Trp53, multiple tumors develop approximately at one year of age. However, while Trp53 knock-out mice predominantly develop osteo- and myosarcomas, a spontaneous loss of both Rad51c and Trp53 in double-mutant mice leads mostly to development of epithelial-derived carcinomas, especially in mammary glands, skin, and skin-associated specialized sebaceous glands. While suggesting a possible role for Trp53 in the Rad51c-mediated tumorigenesis, this study left several questions unaddressed. First, the ability of Rad51c loss to induce tumor formation independently of Trp53 stood as an open question. Second, the mechanisms by which Rad51c might cause malignant transformation remained unclear. Last, there was complete absence of information about the role of RAD51C in the mammary gland. We set out to fill these gaps by generating a skin and skin-associated Rad51c knock-out mouse model. For this purpose, we conditionally deleted Rad51c and/or Trp53 from basal cells of the epidermis and ectodermal-derived glands using Keratin 14 Cre-mediated recombination. With this model, we demonstrated that deletion of Rad51c alone is not sufficient to drive tumorigenesis but impairs the proliferation of sebaceous cells and causes their transdifferentiation into terminally differentiated keratinocytes. In addition, we reported that Rad51c/p53 double mutant mice develop multiple tumors in skin and mammary and sebaceous glands at around six months of age, while Trp53-mutants have a tumor-free survival of 11 months and a lower tumor burden. We also observed that in situ carcinomas are detectable in Rad51c/p53 double mutant mice as early as four months of age, which provided a tool for studying the early phases of tumorigenesis. Notably, we reported that mouse mammary tumors recapitulate several histological features of human RAD51C-associated breast cancers, especially a luminal-like, hormone receptor-positive status. Finally, we described that loss of RAD51C causes chromosomal aberrations in both mouse and human cells, providing a direct translational link between the phenotype observed in the two species.
  • Terhonen, Eeva-Liisa (Helsingin yliopisto, 2015)
    Scots pine (Pinus sylvestris) and Norway spruce (Picea abies) cover large areas in boreal regions with significant economic importance to Finnish forest industry. Approximately 15% of the spruce trees felled in Finland are rotten and thus commercially less valuable. The majority of this conifer wood decay is due to the root and butt rot pathogen Heterobasidion annosum sensu lato. Extensive logging of conifer forests has changed the environment into favouring this pathogen in stands where it originally has been rare. The saprotrophic fungus Phlebiopsis gigantea has for several years been used as a biocontrol agent against H. annosum s.l. in spruce and pine stumps. A major problem is that, although the effectiveness of P. gigantea as a biocontrol agent has empirically been shown, the long-term biological effect of this fungus on other decomposing wood microbiota has not been proven. The first objective of this thesis is to evaluate the impact of the only biocontrol agent used against root and butt rot fungus (H. annosum s.l.) on other resident microflora of Norway spruce stumps. An additional objective is to screen and identify other potentially novel bioagents that can be deployed for the biocontrol of the conifer pathogen. To find out whether the P. gigantea treatment impacts the overall diversity of other non-target stump microbes we used the 454- pyrosequencing approach. Samples were collected from forest sites previously pre-treated with P. gigantea either one, six or 13 years ago. Similarly samples were collected from untreated stumps within the same forest site over the same period of time. The results revealed that initial application of the biocontrol agent influenced the fungal species composition, but the overall fungal diversity was not affected and no statistical differences were observed between treated and non-treated stumps in the mycobiota. The biocontrol treatment significantly decreased the initial bacterial richness in the stumps, but the bacterial community gradually recovered and the negative effect of P. gigantea was attenuated. In parallel to the above studies, I further explored the potential of finding other novel biocontrol agents for use in managing the disease caused by the root rot pathogen Heterobasidion parviporum. This necessitated isolation studies of fungal root endophytes from forestry sites such as pristine mires and drained peatlands where the spread of H. annosum s.l. species have not been commonly reported. A total of 113 isolates of fungal root endophytes were obtained from non-mycorrhizal P. abies roots, which were assigned to 15 different operational taxonomic units (OTUs). Nineteen of the isolates (17%) inhibited the growth of the conifer root rot pathogen in vitro. From these, two isolates were further used to test the potential inhibitory effects during interaction, in vitro, with H. parviporum. Additionally, the metabolites secreted by the selected root endophytes were extracted and the inhibitory effects on these pathogenic fungi were assayed. The secreted metabolites were further chemically analysed using ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-QTOF/MS). These results give new information from the new possibilities to protect the valuable seedling roots from root rot. The results of the thesis provide new information to facilitate better management and protection of these forest sites from H. annosum s.l. as well as support the continued used of P. gigantea for stump pre-treatment in Finnish forests.
  • Gustafsson, Sofia (Finska Vetenskaps-Societeten, 2015)
    In 1748, the Swedish Crown began construction of the fortress Sveaborg outside Helsinki. This was an expensive military investment, but where did the money invested actually end up? Traditionally historians have claimed that the burghers of Helsinki benefitted economically from the construction. However, studies of early-modern Western European fortress construction sites show that local communities did not necessarily benefit directly; the money could just as easily have gone to large entrepreneurs from the central areas. The aim of this study is to identify which geographical areas and social groups sold the construction materials needed to build Sveaborg during the first period of construction in 1748-1756. It also discusses the institutional constraints, whether formal or informal, that limited the suppliers actions and choices. The main source material is the accounts of the Fortification Fund in Helsinki. However, to identify the sellers, we combined this material with local civil and private sources. The accounts rarely provided enough information for identification, and the key-question involved determining where to search for local and private sources. Alfred Weber s theory of location of economic activities claims that the optimal location for production depends on labour and transportation costs. The logistics of Sveaborg were water-based, so the search focused on coastal regions either near the fortress or with unique natural resources. With regard to the institutions, we used the theories of Douglass C. North. The laws of the period limited trade to certain groups in the cities, but equally important were the restrictions on suitable conduct for different estates. However, institutions constantly change, and according to Sten Carlsson, the traditional social system in 18th-century Sweden was already in decline. The results suggest that different economic zones specialising in certain products emerged around the fortress. Suppliers benefitting from Sveaborg could be found not only in the city of Helsinki, but also in the surrounding countryside. Other Finnish cities supplying the fortress include Porvoo, Turku and Loviisa. In Sweden, the main areas involved were Stockholm and Gotland. In contrast to the traditional view, one could definitely not claim that the burghers of Helsinki alone profited from the construction. Although formal constraints should have restricted the participation of other social groups, the informal constraints were obviously more permissive. Unlike in Western Europe, where the states tended to use large entrepreneurs from the central areas, the Swedish Crown used small entrepreneurs from the periphery. The money invested in Sveaborg was spread over large areas and many hands, including peasants, merchants, and noblemen.
  • Louhimo, Riku (Helsingin yliopisto, 2015)
    Cancer is one of the leading causes of death in industrialized nations and its incidence is steadily increasing due to population aging. Cancer constitutes a group of diseases characterized by unwanted cellular growth which results from random genomic alterations and environmental exposure. Diverse genomic and epigenomic alterations separately and jointly regulate gene expression and stimulate and support neoplastic growth. More effective treatment, earlier and more accurate diagnosis, and improved management of cancer are important for public health and well-being. Technological improvements in data measurement, storing and transport capability are transforming cancer research to a data-intensive field. The large increases in the quality and quantity of data for the analysis and interpretation of experiments has made employing computational and statistical tools necessary. Data integration - the combination of different types of measurement data - is a valuable computational tool for cancer research because data integration improves the interpretability of data-driven analytics and can thereby provide novel prognostic markers and drug targets. I have developed two computational data integration tools for large-scale genomic data and a simulator framework for testing a specific type of data integration algorithm. The first computational method, CNAmet, enhances the interpretation of genomic analysis results by integrating three data levels: gene expression, copy-number alteration, and DNA methylation. The second computational method, GOPredict, uses a knowledge discovery approach to prioritize drugs for patient cohorts thereby stratifying patients into potentitally drug-sensitive subgroups. Using the simulator framework, we are able to compare the performance of integration algorithms which integrate gene copy-number data with gene expression data to find putative cancer genes. Our experimental results indicate in simulated, cell line, and primary tumor data that well-performing integration algorithms for gene copy-number and expression data use and process genomic data appropriately. Applying these methods to diffuse large B-cell lymphoma, integrative analysis of copy-number and expression data helps to uncover a gene with putative prognostic utility. Furthermore, analysis of glioblastoma brain cancer data with CNAmet suggests that a number of known cancer genes, including the epidermal growth factor receptor, are highly expressed due to co-occuring alterations in their promoter DNA methylation and copy-number. Finally, integration of publicly available molecular and literature data with GOPredict suggests that treating patients with FGFR inhibitors in breast cancer and CDK inhibitors in ovarian cancer could support standard drug therapies. Collectively, the methods developed here and their application to varied molecular cancer data sets illustrates the benefits of data integration in cancer genomics.
  • Karkman, Antti (Helsingin yliopisto, 2015)
    Increasing microbial resistance against antibiotics is threatening their efficiency in the future and we might be heading back to pre-antibiotic era. Infectious diseases still treatable with antibiotics might soon become life threatening. There is a strong correlation between antibiotic use and antibiotic resistance occurrence. Hotspots for antibiotic resistance are mainly man made such as wastewater treatment plants, animal farms and aquaculture. Aquaculture is the fastest growing food industry in the world and uses antibiotics to treat and prevent fish diseases. The use of antibiotics is linked to increase in antibiotic resistance at the farms. At urban wastewater treatment plants microbes from various sources can mix and exchange genetic material. In addition wastewaters contain antibiotics that can further select for resistant microbes. In this work hundreds of antibiotic resistance genes were quantified with cutting-edge molecular methods. The global patterns of antibiotic resistance gene movement were determined using publicly available metagenomic data. In addition, the famous Baas-Becking hypothesis everything is everywhere, but the environment selects was tested on gene level. Aquaculture increases the amount of antibiotic resistance genes in the farms. The resistance genes persist in the aquaculture sites without a clear selection pressure, however the impact is only local. Urban wastewater treatment plants efficiently removed antibiotic resistance genes from wastewaters. The release of wastewater had only a limited impact on the sediment resistome near the release site. When looking at the metagenomic data, antibiotic resistance genes were found to have different dispersal pattern compared to other genes in the metagenomes. Antibiotic resistance genes can cross taxonomical and geographical barriers with ease, possibly explaining their wide dispersal in the environment and the clinic. These results show that antibiotic resistance is ubiquitous in the environment and the anthropogenic activities affect the incidence of antibiotic resistance.
  • Arovuori, Kyösti (Helsingin yliopisto, 2015)
    This dissertation analyses the effectiveness of the Common Agricultural Policy of the European Union. The motivation of this study arises from the fact that there is a lack of empirical research on the effects of policy instruments on the stated policy objectives. In addition, most of the analysis in the literature has focused on the policy objective to secure farmers incomes and, thus, on the efficiency of income redistribution. The policy effectiveness is defined as the ability of agricultural policy to respond to the stated policy objectives, given the general economic and structural conditions under which the policies operate. In this study, an empirical analysis of the effects of implemented policies and policy reforms on the stated policy objectives in the Common Agricultural Policy of the European Union is conducted. The analysis is carried out at the EU15 level and the time period analysed ranges from 1975 to 2007. In the empirical part, an econometric model utilising panel data for the EU15 countries is built. In the model, the development of the defined policy target variables is explained with policy variables and a set of economic and structural control variables. The results show that policy target variables have, in general, developed in the desired direction. The productivity of agriculture has increased, markets have been stable, self-sufficiency ratios have been achieved and the real term food prices have declined. However, farmers incomes have mostly declined. The implemented agricultural policy reforms have improved the policy effectiveness in general. The main contribution of the implemented reforms has been to the use of resources in agriculture. A policy shift from coupled price support to direct payments has released resources from agriculture to be utilised in other sectors. On the other hand, policy reforms have led to increasing price variation. This is a self-explanatory impact in the sense that administrative price setting was reduced and later on abolished in the policy reforms. According to this study, the impact of agricultural policy on the policy objectives is multifunctional. The implemented policy instruments may also have worked in the opposite direction compared to the targets set. While agricultural policies have clearly contributed to increasing agricultural productivity, they have also absorbed resources into the sector which could have been utilised more efficiently in other sectors. The country-level heterogeneity of economic and agricultural structures has increased especially due to the recent enlargements of the EU from EU15 to EU28. The analysis in this study shows that country-level heterogeneity has a significant impact on the development of policy target variables. Although the implemented policy reforms have made a contribution towards the desired direction and improved the effectiveness of the policies, it is a major challenge for future agricultural policies to effectively tackle the different structures.
  • Soto, Tomás (Helsingin yliopisto, 2015)
    The homogeneous Besov spaces B(s,p,q) and the homogeneous Triebel-Lizorkin spaces F(s,p,q) are two scales of function spaces on the Euclidean space that include the standard and fractional-order Sobolev spaces, and arise naturally in the contexts of e.g. partial differential equations and interpolation theory. In the first article presented in this thesis (joint work with H. Koch, P. Koskela and E. Saksman), we characterize the homeomorphisms of the Euclidean space that induce bounded composition operators on the Besov spaces whose norms are invariant under a scaling change of variable, i.e. where sp equals the dimension of the underlying space, for most indices in the smoothness range 0 < s < 1. We also establish similar results in the setting of certain more general metric measure spaces. In the second article (joint work with M. Bonk and E. Saksman), we present a new characterization of the Triebel-Lizorkin spaces for most indices in the smoothness range 0 < s < 1 in the setting of doubling metric measure spaces, and use the characterization to expand on an earlier result by Koskela, Yang and Zhou concerning the bounded composition operators on the scaling invariant Triebel-Lizorkin spaces. We also establish results concerning general Sobolev-type embeddings and complex interpolation for these function spaces in the generality of doubling metric measure spaces. In the third article, we establish several characterizations of certain larger scales of function spaces on the Euclidean space introduced by Yang and Yuan. One of these characterizations can in particular be applied in the setting of more general metric measure spaces.
  • Luchkina, Natalia V. (Helsingin yliopisto, 2015)
    Activity-dependent synaptic plasticity, and long-term potentiation in particular, represents the predominant model of memory and learning at the cellular level. In addition, synaptic plasticity plays a critical role in the activity-dependent refinement and fine-tuning of neuronal circuits during development by maintaining and stabilising certain synaptic connections and eliminating others. The main goal of this project was to increase our understanding of the molecular mechanisms underlying activity-dependent synaptic plasticity in the developing brain, with particular emphasis on the mechanisms that are specific to early postnatal development. First, we characterise in detail the properties of developmentally restricted neonatal presynaptic long-term potentiation (LTP) in CA1 area of the hippocampus and demonstrate its susceptibility to regulation via protein kinase C (PKC) signalling. Next, we explore the physiological functions of GluA4 subunit-containing AMPA type glutamate receptors, predominantly expressed at developing CA3 CA1 synapses. We show that GluA4 expression is necessary for protein kinase A (PKA)-dependent LTP at immature synapses. Further, the loss of GluA4 expression in parallel with circuit maturation explains the developmental switch in LTP signalling requirements from PKA- to Ca2+/calmodulin-dependent protein kinase II (CaMKII)-dependent. Further, we also explore the role of GluA4 C-terminal interaction partners in synaptic trafficking of GluA4-containing AMPA receptors and its importance for synapse maturation. We confirm a critical role for the membrane proximal region of GluA4 C-terminal domain in trafficking and identify a novel mechanism for activity-dependent synaptic delivery of GluA4 by the extreme C-terminal region. Finally, we show an important role of the GluA4 subunit in strengthening of AMPA receptor-mediated transmission, observed during early postnatal development. In summary, we provide novel information on the pre- and postsynaptic plasticity mechanisms operating at hippocampal CA3 CA1 synapses during the critical period of activity-dependent maturation of glutamatergic neuronal circuitry in rodents. This expands our knowledge on the cellular mechanisms guiding development of synaptic connectivity in the brain. Dysfunction of such mechanisms may play fundamental roles in the underlying pathophysiological causes of various neurodevelopmental disorders.
  • Rautiola, Juhana (2015)
    Renal cell carcinoma (RCC) is the seventh most common cancer in men and the ninth most common in women and it comprises 2-3% of all malignant tumors in adults. In its localized form, the five-year survival is almost 60%. However, in metastatic renal cell carcinoma (mRCC), the five-year survival has historically remained under 10%. Renal cell carcinoma is highly tolerant to traditional chemotherapy and and recent treatment has instead relied on inhibiting the vascular endothelial growth factor (VEGF) and mammalian target of rapamycin (mTOR) signaling pathways. Since 2005, seven new agents have been approved for the treatment of metastatic RCC. Despite the increased knowledge of RCC, the molecular mechanisms behind acquired and intrinsic resistance to treatment remain largely unknown. Our results demonstrate that hypertension is a strong predictor of good prognosis and response to sunitinib treatment. In addition to hypertension we identified neutropenia and thrombocytopenia as predictive markers of good prognosis. A novel, triple biomarker profile consisting of these adverse events strongly predicts improved survival and outcome in mRCC patients. Results from our investigation of sequential treatment suggest that a newer VEGF receptor inhibitor, pazopanib, has clinically meaningful activity in a patient population where mRCC has progressed through sunitinib treatment and in patients who do not tolerate sunitinib. Encouraged by the fuller understanding of tumor biology, we then investigated the role of an endothelial-cell derived growth factor, angiopoietin-2 (Ang2), on patients outcome. High expression of Ang2 was demonstrated to correlate with a high vessel density and to predict a better response to sunitinib-based first-line therapy.
  • Li, Jianyong (Helsingin yliopisto, 2015)
    Assessing the numerical relationships among pollen spectra, human influence and climate is a critical prerequisite for obtaining reliable and robust pollen-based quantitative reconstructions for past climatic variations and human-induced environmental changes. Here, modern pollen data from small river catchments in human-influenced mountainous ecosystems of northern China are collected to investigate the pollen/vegetation/land-use relationships. The effects of human influence index (HII) in comparison with key climate variables on influencing modern pollen distribution are evaluated using a continental-scale dataset. The importance of different climate variables accounting for modern pollen distribution is quantified for identifying the determinant variables that can be reconstructed in different bioclimatic regions of China. Furthermore, pollen-based calibration models for HII and climate parameters are developed and applied to a set of fossil pollen data for quantitative human influence and climate reconstructions. In addition, the bias caused by long-term and intensive human influence on vegetation in quantitative pollen-based palaeoclimatic reconstructions is examined, and pollen-based climate reconstructions are used to validate the reliability of climate model simulations. The results reveal that pollen assemblages from surface sediment samples in the Tuoliang and Qipanshan catchments from northern China are dominated by Artemisia, Chenopodiaceae, Pinus and Selaginella sinensis. Pollen types indicative of human activities such as Cerealia-type are common but not abundant. Artemisia and Chenopodiaceae percentages decline with decreasing altitude, whereas Pinus and S. sinensis percentages increase, suggesting that saccate Pinus pollen and S. sinensis spores can be transported further than non-saccate pollen types and that pollen sorting is taking place over water transportation. There is a positive linear relationship between percentages of Cerealia-type pollen and proportions of farmland, whereas correlation between percentages of trees, shrubs and herbs pollen and proportions of woodland, scrubland and grassland respectively is poor. This can be attributed to the differences in pollen productivity, deposition and dispersal. These findings may provide the basis for a modeling approach to reconstruct past land-cover changes at small catchment scale in northern China. The effects of HII on accounting for modern pollen distribution are smaller than annual precipitation (PANN) or annual average temperature (TANN) in most regions of China, with the exception of east-central China that has a significant pollen–HII relationship. The quantified importance of six climate variables (PANN; TANN; actual/potential evapotranspiration ratio, Alpha; mean temperature of the warmest month, MTWA; mean temperature of the coldest month, MTCO; annual sum of the growing degree days above 5 °C, GDD5) for the geographical distribution of modern pollen data differs among regions in China. Hydrological variables are more important than temperature-related variables in north-western and -eastern China and the Tibetan Plateau, whilst temperature-related variables are dominant in east-central and southern China. This suggests that the critical climate variables which can be optimally reconstructed from pollen data vary regionally in China. These regional differences are common to other large-scale surface pollen datasets and need to be taken into considerations in pollen-based climate reconstructions. The calibration models for HII, PANN and TANN show good statistical performances and are applied to a fossil pollen sequence from Lake Tianchi in east-central China. The inferred HII increases abruptly at 1100 cal. yr BP and remains high until present-day. Correspondingly, the reconstructed PANN shows a sudden decrease since 1100 cal. yr BP, whereas TANN displays an increase. However, other independent palaeoclimatic records mostly contradict the reconstructed climatic trends for the last 1100 years, suggesting that the Tianchi climate reconstructions for this period are biased by the intensification of human influence. This bias probably occurs in other palaeoclimatic reconstructions using pollen data from other regions where human impact on vegetation is also intensive and long-lasting. The calibration models for the summer monsoon precipitation (Pjja) also exhibit good predictive performances and are applied to a set of fossil pollen data from monsoonal China. The Holocene Pjja reconstructions from northern China are stacked to one record, showing that Pjja increases since 9500 cal. yr BP, reaches the maximum at 7000–4000 cal. yr BP, and falls down to present-day. The reconstructions are compared with Pjja simulations from the Kiel Climate Model (KCM). The model output and reconstructions are different for the early-Holocene for which the model suggests higher Pjja. The simulated Pjja changes for the Holocene summer monsoon maximum (HSMM) are lower than the reconstructed changes. The increasing or decreasing Pjja trends before or after the HSMM are more pronounced in the reconstructions than in the model results. Other palaeoclimatic data show substantial Holocene monsoon precipitation changes. This suggests that the KCM underestimates the magnitude of the summer monsoon precipitation changes. The results obtained in this work highlight the necessity for critically assessing the modern pollen/human-influence/climate relationships at various spatial scales and can be used to improve the correctness and robustness of pollen-based climate and human influence reconstructions in China and other regions of the world with large-scale surface pollen datasets. The pollen-based calibration models and their forthcoming applications can be employed to further quantitatively explore the past environmental changes and climatic dynamics and to validate the accuracy of climate model output.
  • Rauhala, Ilona (Helsingin yliopisto, 2015)
    The dissertation consists of five articles, as well as an introductory chapter that explains the background of the study and also draws together the main results of the articles. The study concentrates on the adjectives in the Uralic languages and aims to present the features of the Uralic adjective category in a historical perspective. The data consists mainly of lexical material collected from various etymological and synchronic dictionaries. In addition, several grammatical features of the adjectives are discussed on the basis of grammatical research. The approach is mainly historical-comparative, but also lexical-semantic and morphosyntactic. All of these frameworks are needed to draw a picture of a common category of adjectives in Uralic, the words it may have included, their paths of semantic change, and the morphosyntactic features the category of adjectives may have had. The results of this study complement the earlier research on adjectives, which has been mainly synchronic. The research conducted suggests that by comparing the categories of adjectives in contemporary Uralic languages and by observing the features of the oldest property concept words in Uralic it is possible to reconstruct an adjective category for Proto-Uralic. The results suggest that the category has included words denoting many central semantic groups that indicate an independent lexical category. The adjectives in the contemporary Uralic languages have common morphosyntactic features that probably date back to the Uralic protolanguage. For instance, the Uralic derivative suffix *-(e)TA has mainly functioned as an adjective marker. The study also reviews critically earlier etymologies presented for some property concept words and suggests some new etymologies.
  • Peuralinna, Terhi (Helsingin yliopisto, 2015)
    This thesis project aimed to study the genetic background of common neurodegenerative diseases such as Alzheimer's disease (AD), dementia with Lewy bodies (DLB) and amyotrophic lateral sclerosis (ALS) in the Finnish population. In the first study, we analysed the role of the amyloid precursor protein (APP) and apolipoprotein E (APOE) genes in AD neuropathology. Mutations in the APP gene cause early-onset familial AD and there is some evidence that APP gene variants would play a role in late-onset AD, too. We genotyped more than 50 common variations in the APP gene and sequenced the APP promoter area to detect rare variations in the area, both known and new. In the Finnish elderly population we did not find any of the APP variations to clearly associate with neuropathologically diagnosed AD or with any of the neuropathological features of AD such as cortical β-amyloid, cerebrovascular β-amyloid or neurofibrillary tangles. In addition, the APOEε4 results were updated, using the whole Vantaa85+ cohort. APOE ε4 is currently the strongest known risk factor for late-onset AD. We found a very strong association of APOE ε4 to all neuropathological features of AD. In the second study, we investigated how the common variants in the α-synuclein (SNCA) gene affect different brain pathologies. -synuclein is the main component of the Lewy bodies, which are the pathological hallmarks of PD and DLB, and which are sometimes found also in AD. We genotyped 11 SNPs from the SNCA gene region. We found no association with the cortical β-amyloid and only a faint one with Lewy related pathology. However, we found an association with neurofibrillary tangles. The SNP (rs2572324) associated with a p=0.004 after the Bonferroni correction. The two-locus analysis suggest an independent effect of APOE ε4 (OR=8.67) and rs2572324 (OR=3.36). In the subjects with both risk factors the effect was almost multiplicatively increased (OR=23.3). 35 subjects out of 38 with both risk factors had severe tau-pathology. These results demonstrated the first evidence for a role of genetic variation in SNCA in tau pathology. Moreover, β-amyloid pathology was not associated with the SNCA variants, demonstrating a dissection of genetic effect on the two principal pathological features of AD. In the third study, we performed a whole-genome genotyping using Finnish ALS samples. The Vantaa85+ study subjects were used as controls. This was the first ALS genome-wide association study to find a genome-wide significant association. The location associated with familial ALS was on the chromosome 9p21, which had been noticed before, but the area had been much larger. The area has also been associated with frontotemporal dementia (FTD). Now we managed to define the area with a 42 SNP haplotype. This considerably reduced the area of interest (down to 232 kb) and increased the possibility to find the mutation behind the disease. This haplotype was found in around 40% of the Finnish familial ALS cases and likely explains the high rate of ALS as well as FTD in Finnish population. In the fourth study, we performed a genome-wide association study of DLB in the Vantaa85+ cohort and found two novel areas to be associated with DLB: The c2p21 location with 9 SNP haplotype (p=5.2x10-7) and the c6p21 location with 6 SNP haplotype (p=1.3x10-7). The c6p21 was significant at the genome-wide level. The c2orf21 haplotype has two genes on its area, c2orf72 and SPTBN1. SPTBN1 is the candidate gene since it encodes beta-spectrin, a component of Lewy bodies, while c2orf72 is barely expressed in the brain. The c6p21 associated haplotype block is located in the HLA region and includes HLA-DPB1 and -DPA1 genes. These studies show that the Finnish population is well-suited to study the genetics of neurodegeneration. Our results showed that neuropathologically defined parameters and diagnoses are strongly associated with genetic risk factors, even with relatively low numbers of samples. Hence, phenotypic precision (pathology) is an important element of the statistical power of a study.
  • Mattila, Osmo (Helsingin yliopisto, 2015)
    Forests offer various, sometimes contradictory utilities to their owners and all other users on the global societal and ecological levels. In Finland, meeting the industrial requirements for a stable roundwood supply has defined the forestry service market, as it has been widely supported by the forest owners, the industrial buyers, and the national forest policy. Along with the changes among the owners themselves, demand for forestry services has fragmented. Recently, by introducing the new Forest Act, which gives more freedom for forest owners to choose between management practices, Finnish government has triggered a change that aims at the creation of more market-oriented distribution of forestry services. Based on the concepts of institutional transition at the market level, service-orientation as value-creation logic change, and business model thinking as the unit-level logic, the theoretical objective of the thesis is to define the ongoing renewal of the forestry service market. Using public and private owners as customers and the current forestry service organizations as service providers, the practical aim of this dissertation is to identify potential opportunities and barriers with respect to creating new services in the forestry service market. Methodologically both qualitative interview studies on forestry service organizations (n=22 and n=17) and quantitative multivariate analysis based on survey data with private (n=557) and public (n=139) forest owners are used. According to the results, there is a growing tension in the market environment accelerated by institutional transition: private forest owners are fragmented into multifaceted groups with various needs, while public owners (such as municipalities) are facing versatile user pressures on their publicly owned forests. Therefore, it seems that the traditional roundwood supply approach may no longer match the needs of versatile customer groups. From structural perspective, the established service market dominated by a small number of players is limiting the successful entry of new enterprises. The lack of dynamic middle-sized companies in the Finnish forest sector coupled with difficulties in adopting a more cooperative mind set is proving to be hindrance for renewal of the forestry service market despite the development of information technology, which can facilitate the use of participative methods in forest management and service marketing.
  • Autio, Karoliina (Helsingin yliopisto, 2015)
    Cancer is one of the most common reasons for death in dogs, cats and humans. New therapeutic modalities are necessary to improve disease outcome. One promising approach is oncolytic virotherapy. Until now, the only oncolytic virus evaluated in a clinical trial in veterinary medicine has been canine oncolytic adenovirus, but a clinical trial has been started with oncolytic vaccinia virus (VV) in pet dogs. In cats, oncolytic viruses have not been evaluated in clinical settings. Tumour treatment in dogs and cats could also serve as a model for human cancer therapy. The purpose of the thesis was to evaluate preclinically whether genetically modified oncolytic VV and Semliki Forest virus (SFV) could offer a new treatment modality for dogs and cats. Oncolytic VV was rendered tumour selective by a dual ablation of vaccinia growth factor and thymidine kinase, making it more cancer-specific than previously used VVs in veterinary research. To further increase the efficacy of the VV, an immunostimulatory gene, CD40L, was added to the virus backbone. Avirulent SFV A7(74) has a natural trophism for cancer cells and was not genetically manipulated. Both viruses infected and killed tested cancer cell lines, and VV also infected most of the primary surgical tumour tissues tested. In the nude mouse xenograft model, double deleted VV (vvdd) significantly reduced tumour growth. Interestingly, when in intact monolayers, SCCF1 cells were not killed by VV, but secreted infectious, morphologically abnormal virions. One dog experienced a possible seizure after VV administration, but no other serious adverse events occurred. Vaccinia DNA declined quickly in the blood after virus administration, but was still detectable one week later by qPCR. Only samples taken directly after the VV infusion contained infectious virus, which was not found in any other blood, saliva, urine or faecal samples. Necropsies did not reveal any pathological changes associated with virus administrations. In conclusion, our results show that oncolytic VV and SFV can infect and kill tested canine cancer cell lines, and that VV has antitumoral activity in the mouse xenograft model and it can infect fresh tumour biopsies. In addition, intravenous administration of the viruses did not induce life-threatening adverse events in healthy dogs. These agents thus warrant further evaluation in veterinary medicine.
  • Kulovesi, Pipsa (Helsingin yliopisto, 2015)
    The tear film covers the cornea and conjunctiva providing nutrients to the corneal cells and protecting it from the external environment. Tear film is composed of three intermixed layers. Mucous matrix covers the epithelial cells and is gradually mixed with the aqueous layer which is covered by a thin lipid layer. The lipid layer consists of both polar and nonpolar lipids. Lipids are thought to prevent the collapse of the tear film onto the ocular surface and to retard evaporation from the tears. In this study several in vitro methods were used to study the surface properties and organization of different lipid mixtures resembling those in the tear film lipid layer. Phosphatidylcholine (PC), phosphatidylethanolamine (PE), and freefatty acids (FFA) were used as the polar lipids and cholesterol oleate (CO), triglycerides (TG), and wax esters(WE) were used as the nonpolar lipids. Langmuir film technique was used to examine the behavior of the lipid films during compressions and de-compression. Brewster angle microscopy (BAM) and atomic force microscopy (AFM) were utilized for visualizing the films. Grazing incidence X-ray difraction was used for surface structure studies. The results of experimental studies were compared with coarse-grained molecular dynamics simulations. Custom built system was used to evaluate the evaporation retarding effect of several lipid mixtures containing wax esters. Compression isotherms showed one to two kinks in the compression curve that account for the rearrangement of the lipids in the film for all mixtures studied. Hysteresis was small meaning that these films are very stable and little, if any, solubilization of lipids into the aqueous phase takes place. All lipid films studied were more or less inhomogeneous when viewed with BAM, especially in higher surface pressures. This is most likely caused by the nonpolar lipids aggregating on the lipid film surface. This was also seen in the simulation studies where CO and TG formed circular aggregates on top of the polar lipids. Nonpolar lipids stabilized the films under high compression by arranging so that the lipid film could have a lower surface pressure than would be expected for small surface areas. However, an excess of nonpolar lipids caused the films to be more inhomogenous and to have less stable structure. Lipid mixtures that contained wax esters did not retard evaporation, which suggests that lipids' function in the tear film may have more to do with maintaining a thin tear film and preventing its collapse rather than preventing evaporation. The task of TFLL is still under debate, although it has been regarded to be the evaporation barrier, hindering the movement of water molecules out from the tear fluid. This, however has been questioned in this thesis project. Although some WEs have been shown to retard the evaporation of water, this effect is lost when different lipid species are mixed, as was done in this study where several WE species were mixed with PC, CO, and TG. No evaporation retarding effect was detected for any mixtures studied. Evaporation retarding lipid films must be very tightly packed but this is not possible for TFLL-like films because the tear film lipids must also be suitable for the environment they are in where they must adapt to fast changes in surface pressure and must also be fluid. This cannot be achieved by the films that have been shown to retard evaporation as these films are rather stiff and cannot be compressed.
  • Kupari, Jussi (Helsingin yliopisto, 2015)
    The Glial cell line-derived neurotrophic factor (GDNF) family ligands, which include GDNF, neurturin (NRTN), persephin (PSPN) and artemin (ARTN), signal through a glycosyl phosphatidyl inositol (GPI)-linked cognate-receptor (GFRα1-4) and the transmembrane receptor tyrosine kinase receptor RET. The members of the GDNF family play a particularly important role in the development of the peripheral nervous system (PNS). In the autonomic nervous system, GDNF and NRTN regulate important steps in the development of the enteric and parasympathetic nervous systems from migration and proliferation to soma size and target innervation, whereas ARTN takes part in the early phases of sympathetic nervous system development. In the sensory system, GFRα2 ¬ the co-receptor of NRTN has been shown to mediate trophic signaling for nonpeptidergic nociceptive neurons, and is also required for their innervation of the glabrous epidermis. However, several aspects of the role of GFRα2-signaling in normal PNS development and function remain poorly understood. Therefore, the aims of this study were to elucidate (1) the role of GFRα2-signaling in the development of parasympathetic neurons; (2) the role of GFRα2-signaling in two classes of somatosensory mechanoreceptor neurons and their target innervation; and (3) the role of GFRα2-signaling in the cholinergic innervation of the gastric mucosa and the role of this innervation in gastric secretion. We discovered that programmed cell death (PCD) is a normal part of parasympathetic neuron development in mice. GFRα2-signaling was found to regulate parasympathetic neuron survival in pancreatic and submandibular ganglia during late embryonic development; lack of GFRα2-mediated signaling resulted in the loss of intrapancreatic neurons through PCD. In argreement with previous studies, apoptosis in the ENS was found to be rare, and was not increased in the absence of GFRα2, implying that the normal number of enteric neurons is not determined by PCD. In the dorsal root ganglia (DRGs), we found that GFRα2 regulates the cell size, but not the peripheral innervation of hair follicles in both the large early-RET RA Aβ-class low threshold mechanoreceptors (LTMRs) and in the small C-LTMRs. In contrast, GFRα2 was found to regulate both the cell size and the epidermal innervation in the small Mrgprd+ C-nociceptors. We also found evidence that the RA Aβ-LTMRs downregulate GFRα2-expression at some point after birth, suggesting a possible switch in neurotrophic signaling pathways. In the enteric nervous system, we demonstrated that GFRα2-signaling via NRTN is required for cholinergic innervation of the gastric mucosa. Interestingly, this innervation was found to be unnecessary for maintaining the gastric mucosa and for gastrin secretion and basal acid secretion. Even though vagally-stimulated secretion is lost in the GFRα2-KO mice, their ability to secrete acid in response to direct parietal cell stimulation remains in the absence of gastric mucosal innervation.
  • Venhoranta, Heli (Helsingin yliopisto, 2015)
    Dairy cattle breeding programs rely heavily on artificial insemination (AI). The AI enables intense selection for desired traits, but the use of few elite bulls carries a risk that inherited defects may proliferate in the population. The emergence of inherited defects is a recurrent issue in cattle breeding and rapid identification and management of genetic defects are crucial for preventing economic losses and maintaining good animal welfare. Modern molecular genetics techniques enable efficient detection of causative mutations, information which is needed for DNA testing. In this study, we investigated three inherited congenital bovine defects and located causative mutations for these diseases. 1. A microdeletion in PEG3 domain causes intrauterine growth restriction and stillbirths to almost half of the pregnancies sired by a single Ayrshire AI bull. The deletion, when inherited from the sire, is semi-lethal but the female mutation carriers can breed normally. The deletion truncates the 3 end of the non-coding maternally imprinted MIMT1 transcript and also causes expression changes in other genes of the domain. The expression changes of hundreds of genes in the foetal side of the placenta were also demonstrated. 2. Inherited gonadal hypoplasia with incomplete penetrance in Northern Finncattle and Swedish Mountain breed is associated with homozygosity for the Cs29 allele. This allele is an ectopic segment that is duplicated and translocated from chromosome 6 to 29 and encompasses the KIT gene. Cs29 allele is associated with colour sidedness in various cattle breeds, which coheres with the results that gonadal hypoplasia is connected with white coat colour. The KIT gene is known to regulate the migration of the germ cells and precursors of melanocytes. 3. A new recessive developmental disorder defined as PIRM (ptosis, intellectual disability, retarded growth and mortality) in Ayrshire breed was associated with single base substitution in the UBE3B gene. Mutation causes in-frame exon skipping, resulting in an altered protein lacking 40 amino acids, which likely comprises protein function. In humans, mutations in the UBE3B gene are associated with Kaufman oculocerebrofacial syndrome, with similar pathological effects as for PIRM syndrome. The UBE3B mutation may be connected with the AH1 haplotype, which is associated with reduced fertility and has a carrier frequency of 26.1% in the North American Ayrshire population. The mutations related to these three defects or syndromes can now be easily tested for. The results can be used to avoid risky matings, cull carriers and provide a veterinary diagnostic. The expression analyses proved that the mutations in Ayrshires also affected the RNA expression of the respective mutated gene or even hundreds of other genes. The described genotype-phenotype associations can be used as a basis for approaches to locate quantitative trait loci in cattle or provide new insights into developmental biology and inform translational research across species.